Androgenic steroids induce pathologic scarring in a preclinical porcine model via dysfunctional extracellular matrix deposition.

Hypertrophic scarring is a major source of morbidity. Sex hormones are not classically considered modulators of scarring. However, based on increased frequency of hypertrophic scarring in patients on testosterone, we hypothesized that androgenic steroids induce abnormal scarring and developed a preclinical porcine model to explore these effects. Mini-swine underwent castration, received no testosterone (noT) or biweekly testosterone therapy (+T), and underwent excisional wounding. To create a delayed wound healing model, a subset of wounds were re-excised at 2 weeks. Scars from postoperative day 42 (POD42) and delayed wounds (POD28) were harvested 6 weeks after initial wounding for analysis via histology, bulk RNA-seq, and mechanical testing. Histologic analysis of scars from +T animals showed increased mean fibrosis area (16 mm2noT, 28 mm2+T; p = .007) and thickness (0.246 mm2noT, 0.406 mm2+T; p < .001) compared to noT. XX+T and XY+T scars had greater tensile burst strength (p = .024 and p = .013, respectively) compared to noT swine. Color deconvolution analysis revealed greater deposition of type I and type III collagen as well as increased collagen type I:III ratio in +T scars. Dermatopathologist histology scoring showed that +T exposure was associated with worse overall scarring (p < .05). Gene ontology analysis found that testosterone exposure was associated with upregulation of cellular metabolism and immune response gene sets, while testosterone upregulated pathways related to keratinization and laminin formation on pathway analysis. In conclusion, we developed a preclinical porcine model to study the effects of the sex hormone testosterone on scarring. Testosterone induces increased scar tissue deposition and appears to increase physical strength of scars via supraphysiologic deposition of collagen and other ECM factors. The increased burst strength seen in both XX and XY animals suggests that hormone administration has a strong influence on scar mechanical properties independent of chromosomal sex. Anti-androgen topical therapies may be a promising future area of research.

Screening oral drugs for their interactions with the intestinal transportome via porcine tissue explants and machine learning.

In vitro systems that accurately model in vivo conditions in the gastrointestinal tract may aid the development of oral drugs with greater bioavailability. Here we show that the interaction profiles between drugs and intestinal drug transporters can be obtained by modulating transporter expression in intact porcine tissue explants via the ultrasound-mediated delivery of small interfering RNAs and that the interaction profiles can be classified via a random forest model trained on the drug-transporter relationships. For 24 drugs with well-characterized drug-transporter interactions, the model achieved 100% concordance. For 28 clinical drugs and 22 investigational drugs, the model identified 58 unknown drug-transporter interactions, 7 of which (out of 8 tested) corresponded to drug-pharmacokinetic measurements in mice. We also validated the model's predictions for interactions between doxycycline and four drugs (warfarin, tacrolimus, digoxin and levetiracetam) through an ex vivo perfusion assay and the analysis of pharmacologic data from patients. Screening drugs for their interactions with the intestinal transportome via tissue explants and machine learning may help to expedite drug development and the evaluation of drug safety.

A Novel Mouse Model for Investigating the Effects of Gender-affirming Hormone Therapy on Surgical Healing.

Wound healing problems are a major cause of morbidity for gender-affirming surgery (GAS) patients. Prior studies have shown sex differences in wound healing may exist. We hypothesized exogenous testosterone supplementation may impair post-GAS wound healing and developed a model to investigate this phenomenon. Mice were randomized by hormone regimen and gonadectomy (OVX). Gonadectomy or sham occurred on day 0 and mice were assigned to no testosterone (-T), mono- or bi-weekly (T/2T) testosterone groups. Dorsal splinted wounding occurred on day 14 and harvest on day 21. Serum testosterone levels were quantified with mass spectrometry. Tissue underwent analysis with planimetry, qPCR, ELISA, and immunofluorescence. Mean testosterone trough levels for bi-weekly regimen were higher compared to mono-weekly (397 versus 272 ng/dL; P = 0.027). At POD5, 2T injections led to 24.9% and 24.7% increases in mean wound size relative to SHAM and OVX/-T, respectively (P = 0.004; 0.001). Wounds in OVX/+2T mice demonstrated increased gene expression for inflammatory cytokines and macrophage marker F4/80 (P < 0.05). ELISA confirmed elevated wound TNFα levels (P < 0.05). Quantitative multiplex immunofluorescence with F4/80/NOS2/ARG1 showed significant increases in macrophage prevalence in OVX/+2T (P < 0.05). We developed a novel model of GAS hormonal milieu to study effects of exogenous testosterone on wound healing. Optimized twice-weekly dosing yielded serum levels comparable to clinical therapy. We showed exogenous testosterone administered to XX/OVX mice significantly impairs wound healing. A hyperinflammatory wound environment results in increased macrophage proliferation and elevated cytokines. Future efforts are directed toward mechanistic investigation and clinical validation.

A microneedle platform for buccal macromolecule delivery.

Alternative means for drug delivery are needed to facilitate drug adherence and administration. Microneedles (MNs) have been previously investigated transdermally for drug delivery. To date, drug loading into MNs has been limited by drug solubility in the polymeric blend. We designed a highly drug-loaded MN patch to deliver macromolecules and applied it to the buccal area, which allows for faster delivery than the skin. We successfully delivered 1-mg payloads of human insulin and human growth hormone to the buccal cavity of swine within 30 s. In addition, we conducted a trial in 100 healthy volunteers to assess potential discomfort associated with MNs when applied in the oral cavity, identifying the hard palate as the preferred application site. We envisage that MN patches applied on buccal surfaces could increase medication adherence and facilitate the painless delivery of biologics and other drugs to many, especially for the pediatric and elderly populations.

Development of a long-acting direct-acting antiviral system for hepatitis C virus treatment in swine.

Chronic hepatitis C virus (HCV) infection is a leading cause of cirrhosis worldwide and kills more Americans than 59 other infections, including HIV and tuberculosis, combined. While direct-acting antiviral (DAA) treatments are effective, limited uptake of therapy, particularly in high-risk groups, remains a substantial barrier to eliminating HCV. We developed a long-acting DAA system (LA-DAAS) capable of prolonged dosing and explored its cost-effectiveness. We designed a retrievable coil-shaped LA-DAAS compatible with nasogastric tube administration and the capacity to encapsulate and release gram levels of drugs while resident in the stomach. We formulated DAAs in drug-polymer pills and studied the release kinetics for 1 mo in vitro and in vivo in a swine model. The LA-DAAS was equipped with ethanol and temperature sensors linked via Bluetooth to a phone application to provide patient engagement. We then performed a cost-effectiveness analysis comparing LA-DAAS to DAA alone in various patient groups, including people who inject drugs. Tunable release kinetics of DAAs was enabled for 1 mo with drug-polymer pills in vitro, and the LA-DAAS safely and successfully provided at least month-long release of sofosbuvir in vivo. Temperature and alcohol sensors could interface with external sources for at least 1 mo. The LA-DAAS was cost-effective compared to DAA therapy alone in all groups considered (base case incremental cost-effectiveness ratio $39,800). We believe that the LA-DAA system can provide a cost-effective and patient-centric method for HCV treatment, including in high-risk populations who are currently undertreated.

Light-degradable hydrogels as dynamic triggers for gastrointestinal applications.

Triggerable materials capable of being degraded by selective stimuli stand to transform our capacity to precisely control biomedical device activity and performance while reducing the need for invasive interventions. Here, we describe the development of a modular and tunable light-triggerable hydrogel system capable of interfacing with implantable devices. We apply these materials to two applications in the gastrointestinal (GI) tract: a bariatric balloon and an esophageal stent. We demonstrate biocompatibility and on-demand triggering of the material in vitro, ex vivo, and in vivo. Moreover, we characterize performance of the system in a porcine large animal model with an accompanying ingestible LED. Light-triggerable hydrogels have the potential to be applied broadly throughout the GI tract and other anatomic areas. By demonstrating the first use of light-degradable hydrogels in vivo, we provide biomedical engineers and clinicians with a previously unavailable, safe, dynamically deliverable, and precise tool to design dynamically actuated implantable devices.

A luminal unfolding microneedle injector for oral delivery of macromolecules.

Insulin and other injectable biologic drugs have transformed the treatment of patients suffering from diabetes1,2, yet patients and healthcare providers often prefer to use and prescribe less effective orally dosed medications3-5. Compared with subcutaneously administered drugs, oral formulations create less patient discomfort4, show greater chemical stability at high temperatures6, and do not generate biohazardous needle waste7. An oral dosage form for biologic medications is ideal; however, macromolecule drugs are not readily absorbed into the bloodstream through the gastrointestinal tract8. We developed an ingestible capsule, termed the luminal unfolding microneedle injector, which allows for the oral delivery of biologic drugs by rapidly propelling dissolvable drug-loaded microneedles into intestinal tissue using a set of unfolding arms. During ex vivo human and in vivo swine studies, the device consistently delivered the microneedles to the tissue without causing complete thickness perforations. Using insulin as a model drug, we showed that, when actuated, the luminal unfolding microneedle injector provided a faster pharmacokinetic uptake profile and a systemic uptake >10% of that of a subcutaneous injection over a 4-h sampling period. With the ability to load a multitude of microneedle formulations, the device can serve as a platform to orally deliver therapeutic doses of macromolecule drugs.

A gastric resident drug delivery system for prolonged gram-level dosing of tuberculosis treatment.

Multigram drug depot systems for extended drug release could transform our capacity to effectively treat patients across a myriad of diseases. For example, tuberculosis (TB) requires multimonth courses of daily multigram doses for treatment. To address the challenge of prolonged dosing for regimens requiring multigram drug dosing, we developed a gastric resident system delivered through the nasogastric route that was capable of safely encapsulating and releasing grams of antibiotics over a period of weeks. Initial preclinical safety and drug release were demonstrated in a swine model with a panel of TB antibiotics. We anticipate multiple applications in the field of infectious diseases, as well as for other indications where multigram depots could impart meaningful benefits to patients, helping maximize adherence to their medication.

3D-Printed Gastric Resident Electronics.

Long-term implantation of biomedical electronics into the human body enables advanced diagnostic and therapeutic functionalities. However, most long-term resident electronics devices require invasive procedures for implantation as well as a specialized receiver for communication. Here, a gastric resident electronic (GRE) system that leverages the anatomical space offered by the gastric environment to enable residence of an orally delivered platform of such devices within the human body is presented. The GRE is capable of directly interfacing with portable consumer personal electronics through Bluetooth, a widely adopted wireless protocol. In contrast to the passive day-long gastric residence achieved with prior ingestible electronics, advancement in multimaterial prototyping enables the GRE to reside in the hostile gastric environment for a maximum of 36 d and maintain ≈15 d of wireless electronics communications as evidenced by the studies in a porcine model. Indeed, the synergistic integration of reconfigurable gastric-residence structure, drug release modules, and wireless electronics could ultimately enable the next-generation remote diagnostic and automated therapeutic strategies.